During atherogenesis, vascular smooth muscle cells are thought to undergo conversion from a contractile to a "synthetic" phenotype, acquiring the ability to proliferate. Our studies over the past several years have shown that the ECM and integrins have essential roles in activating the G1 phase cyclin-dependent kinases (cdks), the critical regulators of cell proliferation. New data described here shows that CD44, an adhesion receptor that binds to hyaluronic acid (HA), regulates integrin-dependent signaling events in aortic smooth muscle cells and fibroblasts, controlling both the actin cytoskeleton and G1 phase cell cycle progression. These results validate a long-standing assumption about the potential interaction between integrins and CD44. Genetic analysis in CD44-null mice suggest that the CD44-integrin interaction may explain, at least in part, the pro-atherosclerotic effect of CD44. Three aims are now proposed to study the interactions between integrins and CD44 on actin organization, G1 phase cell cycle progression, and aortic smooth muscle cell proliferation. Aim 1 will extend preliminary results showing that CD44 inhibits integrin-dependent cell spreading and formation of actin stress fibres. We will characterize potential physical interactions between CD44 and integrins and study the mechanism by which CD44 affects integrin function. Aim 2 extends preliminary data showing that HA and CD44 regulate G1 phase cell cycle progression. We will map the molecular basis of the regulation and determine how HA-CD44 affects the signaling pathways regulating G1 phase progression. Aim 3 examines the effect of CD44 on cell proliferation in vivo by mating CD44-null and CD44/ApoE-double null mice to a cyclin A promoter-eGFP transgenic mouse. Analysis of eGFP fluorescence in isolated aortae from the progeny will allow us to determine the effect of CD44 on normal smooth muscle cell proliferation, as well as on the stimulated smooth muscle cell proliferation thought to occur during atherosclerosis and catheter-induced injury.